This invention relates to vascular endothelial growth factor. More specifically, this invention relates to oligonucleotides specific for vascular endothelial growth factor nucleic acid and useful treatment of disorders that are associated with neovascularization and angiogenesis, such as psoriasis.
Neovascular diseases of the retina such as diabetic retinopathy, retinopathy of prematurity, and age-related macular degeneration are a major cause of blindness in the United States and the world, yet the biochemical events responsible for these processes have not been fully elucidated.
Diabetic retinopathy is the leading cause of blindness among working age adults (20-64) in the United States (Foster in Harrison's Principles of Internal Medicine (Isselbacher et al., eds.) McGraw-Hill, Inc., New York (1994) pp. 1994-1995). During the course of diabetes mellitus, the retinal vessels undergo changes that result in not only leaky vessels but also vessel drop out resulting in retinal hypoxia. The effects of these complications are hemorrhaging, "cotton wool" spots, retinal infarcts, and neovascularization of the retina resulting in bleeding and retinal detachment. If left untreated, there is a 60% chance of visual loss. Classic treatment for proliferative diabetic retinopathy is panretinal laser photocoagulation (PRP). However, complications can occur from panretinal laser photocoagulation such as foveal burns, hemorrhaging, retinal detachment, and choroidal vessel growth. Furthermore, other untoward effects of this treatment are decreased peripheral vision, decreased night vision, and changes in color perception (Am J. Ophthalmol. (1976) 81:383-396; Ophthalmol (1991) 98:741-840). Thus, there is a need for a more effective treatment for diabetic retinopathy.
Retinopathy of prematurity (ROP) is a common cause of blindness in children in the United States (Pierce et al. (1994) Int. Ophth. Clinics 34:121-148). Premature babies are exposed to hyperoxic conditions after birth even without supplemental oxygen because the partial pressure of oxygen in utero is much lower than what is achieved when breathing normal room air. This relative hyperoxia is necessary for their survival yet can result in ROP. The blood vessels of the retina cease to develop into the peripheral retina resulting in ischemia and localized hypoxic conditions as the metabolic demands of the developing retina increase. The resulting hypoxia stimulates the subsequent neovascularization of the retina. This neovascularization usually regresses but can lead to irreversible vision loss. There are at least 10,000 new cases per year with a worldwide estimate of 10 million total cases. At present, there is no effective cure for ROP. Two therapeutic methods, cryotherapy and laser therapy, have been used but are not completely effective and themselves cause damage to the eye, resulting in a reduction of vision (Pierce et al. (1994) Int. Ophth. Clinics 34:121-148). Many other antiangiogenic compounds have been tested, but no inhibition in retinal neovascularization has been reported (Smith et al. (1994) Invest. Ophthamol Vis. Sci. 35:1442; Foley et al. (1994) Invest. Ophthamol Vis. Sci. 35:1442). Thus, there is a need for an effective treatment for ROP.
Age related macular degeneration is one of the leading causes of blindness in older adults in the United States, and may account for up to 30% of all bilateral blindness among Caucasian Americans (Anonymous (1994) Prevent Blindness America). This disease is characterized by loss of central vision, usually in both eyes, due to damage to retinal pigment epithelial cells which provide physiological support to the light sensitive photoreceptor cells of the retina. In most cases there is currently no effective treatment. In approximately 20% of exudative cases that are diagnosed early, laser treatment can prevent further loss of vision; however, this effect is temporary (Bressler et al., Principles and Practices of Ophthalmology (eds. Albert and Jakobiac), W. B. Saunders Co., Philadelphia, Pa.) (1994) Vol. 2 pp. 834-852). Thus, there is a need for a more effective and permanent treatment for age related macular degeneration.
Ocular neovascularization is also the underlying pathology in sickle cell retinopathy, neovascular glaucoma, retinal vein occlusion, and other hypoxic diseases. These eye diseases as well as other pathological states associated with neovascularization (i.e., tumor growth, wound healing) appear to have hypoxia as a common factor (Knighton et al. (1983) Science 221:1283-1285; Folkman et al. (1987) Science 235:442-446; Klagsbrun et al. (1991) Ann Rev. Physiol 53:217-239; Miller et al. (1993) Principles and Practice of Ophthalmology, W. B. Saunders, Philadelphia, pp. 760; and Aiello et al. (1994) New Eng. J. Med. 331:1480-1487). Moreover, retinal neovascularization has been hypothesized to be the result of a "vasoformative factor" which is released by the retina in response to hypoxia (Michaelson (1948) Trans. Ophthamol. Soc. U. K. 68:137-180; and Ashton et al. (1954) Br. J. Ophthalmol. 38:397-432). Recent experimental data show a high correlation between vascular endothelial growth factor expression and retinal neovascularization (Aiello et al. (1994) New Eng. J. Med. 331:1480-1487). Furthermore, elevated levels of vascular endothelial growth factor have recently been found in vitreous from patients with diabetes (Aiello et al., ibid.). Thus, this cytokine/growth factor may play an important role in neovascularization-related disease.
Vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) is an endothelial cell-specific mitogen which supports angiogenesis in wound healing and development. In addition, VEGF has recently been shown to be stimulated by hypoxia and required for tumor angiogenesis (Senger et al. (1986) Cancer 46:5629-5632; Kim et al. (1993) Nature 362:841-844; Schweiki et al. (1992) Nature 359:843-845; Plate et al. (1992) Nature 359:845-848). It is a 34-43 kD (with the predominant species at about 45 kD) dimeric, disulfide-linked glycoprotein synthesized and secreted by a variety of tumor and normal cells. In addition, cultured human retinal cells such as pigment epithelial cells and pericytes have been demonstrated to secrete VEGF and to increase VEGF gene expression in response to hypoxia (Adamis et al. (1993) Biochem. Biophys. Res. Commun. 193:631-638; Plouet et al. (1992) Invest. Ophthamol. Vis. Sci. 34:900; Adamis et al. (1993) Invest. Ophthamol. Vis. Sci. 34:1440; Aiello et al. (1994) Invest. Ophthamol. Vis. Sci. 35:1868; Simorre-Pinatel et al. (1994) Invest Ophthamol. Vis. Sci. 35:3393-3400). In contrast, VEGF in normal tissues is relatively low.
VEGF has also been shown to play a major role in other diseases associated with the aberrant angiogenesis, including tumor development and skin disorders. Conditions associated with such irregularities include cancer, rheumatoid arthritis, the bullous diseases (including bullous pemphigoid, dermatitis herpetiformis, and erythema multiforme), and psoriasis.
Psoriasis is a chronic skin disorder that affects one in fifty people world wide and over five million people in the United States. Approximately 150,000 to 250,000 new cases of psoriasis are diagnosed each year. Ten percent of people with psoriasis develop psoriatic arthritis. The most common form of the disease is called plaque psoriasis or psoriasis vulgaris. Other forms are pustular, guttate, inverse, and erythrodermic psoriasis.
The cause of psoriasis is unknown. The skin lesions of psoriasis vulgaris are in part a result of an excessive rapid growth and turnover of keratinocytes. Current types of treatment achieve some temporary relief (e.g., steroids, anthralin, calcipotriene, coal tar with or without light therapy, psoralen with UVA treatment, methotrexate, retinoids with or without UV light, and cyclosporin A). However, harmful side effects of these treatments exist which vary in degree of health hazard, and some must be carefully monitored by a physician. Furthermore most of these treatments result in a recurrence of the psoriasic symptoms.
Ballaun et al. (J Invest. Dermatol. (1995) 104:7-10) have shown that the three major splice forms of VEGF are produced by human epidermal keratinocytes and can be detected in the supernatant of cell cultures. In addition, VEGF upregulation has been observed in bullous pemphigoid, dermatitis herpetiformis, and erythema multiforme (Brown et al. (1995) J. Invest. Dermatol. 104:744-749). Furthermore, psoriasic lesions express elevated levels of transforming growth factor alpha (TGF.alpha.), a known inducer of VEGF in cell culture (Detmar et al. (J. Invest Dermatol. (1995) 105:44-50), and VEGF levels of normal epidermal keratinocytes can be induced in response to elevated levels of TGF.alpha..
Thus, VEGF appears to play a principle role in many pathological states and processes related to angiogenesis and neovascularization. Regulation of VEGF expression in tissues affected by the various conditions described above could therefore be key in treatment or preventative therapies associated with such disorders.
New chemotherapeutic agents termed "antisense oligonucleotides" have been developed which are capable of modulating cellular and foreign gene expression (see, Zamecnik et al. (1978) Proc. Natl. Acad. Sci. (USA) 75:280-284). Without being limited to any theory or mechanism, it is generally believed that the activity of antisense oligonucleotides depends on the binding of the oligonucleotide to the target nucleic acid (e.g. to at least a portion of a genomic region, gene or mRNA transcript thereof), thus disrupting the function of the target, either by hybridization arrest or by destruction of target RNA by RNase H (the ability to activate RNase H when hybridized to RNA).
VEGF-specific antisense oligonucleotides have been developed (Uchida et al. (1995) Antisense Res. & Dev. 5(1):87 (Abstract OP-10); Nomura et al., (1995) Antisense Res. & Dev. 5(1):91 (Abstract OP-18)), although none have been demonstrated to reverse neovascularization or angiogenesis. Thus, a need still remains for the development of oligonucleotides that are capable of reducing VEGF expression, and ultimately, of inhibiting the onset of diseases and disorders associated with the expression of VEGF.